The studies proposed here are designed to elucidate the control and function(s) of some enzymes involved in the regulation of the deoxythymidylate pool in normal proliferating, differentiated and malignant tissues in man. The basic hypothesis is that a progressive shift occurs in the isozyme pattern during carcinogenesis that results in TdR kinase activity biochemically and/or antigenically distinct from that of normal proliferating cells. The various forms of TdR kinase will be purified from human tissues and characterized by their biochemical, physico-chemical and antigenic properties. Purification procedures will include classical, preparative electrophoretic and isoelectric focusing techniques as well as affinity chromatography (TdR analog or antibody as ligand). The latter will be used for rapid purification of enzymes from small quantities of tissues. Antibodies to purified enzymes will be used in studies on antigenic relationships of kinases from diverse sources and as ligands for immunosorbents, Sedimentation profiles in the presence and absence of effectors and cell regulatory compounds, e.g., cyclic AMP and polyamines, and binding studies of the latter compounds (equilibrium dialysis) will be used to assess if the aggregated state(s) of TdR kinase could be a means of controlling its activity. The spectrum of isozyme patterns concomitant with high-risk for cancer will be determined in tissues and fibroblasts from patients with Gardner's syndrome. Essentially the same protocol will be followed for TdR phosphorylase. Kinetic parameters of the two enzymes determined at limiting and at excess TdR concentrations will be used to assess their relative roles in competing for TdR. Evaluation of all these parameters as well as intracellular pool sizes of nucleosides and nucleotides should aid in understanding the cellular function and regulation of TdR kinase and perhaps confirm that the initial observations of differences between colon and tumor are not unique for this tissue pair and that tumor-specific TdR kinase isozyme(s) could play a critical role in malignant growth in general.